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Gut anaerobes protect against pathogen invasion Intestinal infections are a common problem for young animals. One explanation is that the protective gut microbiota is not fully established in infants. How the microbiota might protect against pathogens is unclear. Kim et al. found that members of the group of strictly anaerobic, spore-forming bacteria known as clostridia protect neonatal mice against diarrhea-causing pathogens. The protective effect is enhanced by giving mice the metabolite succinate in drinking water. Succinate favors colonization of the neonatal gut by cluster IV and XIVa clostridia and concomitantly excludes Salmonella typhimurium. Science , this issue p. 315

Introduction : tRNA-derived fragments (tRFs) have been postulated to play a significant role in human disease and specifically cancer. tRFs can function as small non-coding RNAs, expressed more highly in conditions of oxidative stress and tissue damage, and are postulated to have a regulatory function. Previous work has identified overexpression of tRNAs in multiple myeloma (MM), but the expression of tRFs in MM has not yet been characterized. In this descriptive study, we set out to apply modern next-generation sequencing tools to describe tRNA and tRF expression in MM. Methods: Bone marrow aspirate samples from 24 patients, 8 with monoclonal gammopathy of undetermined significance (MGUS) or smoldering MM (SMM), 8 with newly diagnosed MM (NDMM), and 8 with relapsed/refractory MM (RRMM) were selected. Total RNA fraction from mononuclear cell fractions was extracted and used to prepare tRNA sequencing libraries which generated data of different small RNA families such as tRNA, tRF, and 5S rRNA in one library. Results: We found a trend toward increased total expression of tRFs in NDMM and RRMM patient samples. Analysis of specific classes of tRFs revealed differential expression along the course of disease progression, with specific tRFs increasing in expression from MGUS/SMM to RRMM. Specifically, tRFs from tRNAHis showed a significant increase in expression from MGUS/SMM to NDMM (p Conclusions : These preliminary results indicate that tRNA fragment expression has the potential to serve as an objective, quantitative measure of clinical MM progression. While the function of these small RNA species remains unclear, specific cleavage patterns suggest that tRF expression in MM may be regulated during disease progression. Future work should explore potential for miRNA-like function of tRFs in MM, and ongoing work to be presented at the meeting will further explore tRF abundance as well as post-transcriptional modifications of tRNAs which may provide a mechanistic understanding of patterns of tRNA fragmentation. Citation Format: Alexandra E. Rojek, Christopher D. Katanski, Andrew Stefka, Benjamin A. Derman, Andrzej Jakubowiak, Tao Pan. tRNA expression and tRFs in multiple myeloma: Progression from monoclonal gammopathies to relapsed/refractory disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2394.

Environmentally induced alterations in the commensal microbiota have been implicated in the increasing prevalence of food allergy. We show here that sensitization to a food allergen is increased in mice that have been treated with antibiotics or are devoid of a commensal microbiota. By selectively colonizing gnotobiotic mice, we demonstrate that the allergy-protective capacity is conferred by a Clostridia-containing microbiota. Microarray analysis of intestinal epithelial cells from gnotobiotic mice revealed a previously unidentified mechanism by which Clostridia regulate innate lymphoid cell function and intestinal epithelial permeability to protect against allergen sensitization. Our findings will inform the development of novel approaches to prevent or treat food allergy based on modulating the composition of the intestinal microbiota.

Tregs play a crucial role in the maintenance of intestinal immune homeostasis. However, significant numbers of Foxp3+ Tregs accumulate in the inflamed lesions in experimental colitis and in IBD patients. Treg production of the proinflammatory cytokines IFN-γ and/or IL-17 may arguably explain their ineffectiveness in suppressing intestinal inflammation. However, it remains unknown whether iTreg and tTreg produce proinflammatory cytokines and how TLR signaling regulates this process. Here, we found that Foxp3+Tregs were increased in the intestines of B6.TLR4−/− and B6.IL-10−/− mice when compared with WT B6 mice. TLR4−/− and IL-10−/− resulted in more Tregs within inflamed intestines. The majority of Foxp3+ Tregs in the spleen was Helios+Nrp1+, whereas most Foxp3+ Tregs in the intestinal LP were Helios−Nrp1−. More Helios+Nrp1+ Tregs expressed IFN-γ and/or IL-17 than did Helios−Nrp1− Tregs in the spleen and intestine, which was increased with TLR4−/−. TLR4 signaling in T cells and APCs inhibited Foxp3+ induction via MyD88-dependent, TRIF-independent pathways, which was negatively regulated by SOCS3. Collectively, these data demonstrate Helios+Nrp1+ tTregs and Helios−Nrp1− iTregs produce proinflammatory cytokines in the intestines during inflammation, which was regulated by TLR4 signaling.

Dietary intervention with extensively hydrolyzed casein formula supplemented with Lactobacillus rhamnosus GG (EHCF+LGG) accelerates tolerance acquisition in infants with cow's milk allergy (CMA). We examined whether this effect is attributable, at least in part, to an influence on the gut microbiota. Fecal samples from healthy controls (n=20) and from CMA infants (n=19) before and after treatment with EHCF with (n=12) and without (n=7) supplementation with LGG were compared by 16S rRNA-based operational taxonomic unit clustering and oligotyping. Differential feature selection and generalized linear model fitting revealed that the CMA infants have a diverse gut microbial community structure dominated by Lachnospiraceae (20.5±9.7%) and Ruminococcaceae (16.2±9.1%). Blautia, Roseburia and Coprococcus were significantly enriched following treatment with EHCF and LGG, but only one genus, Oscillospira, was significantly different between infants that became tolerant and those that remained allergic. However, most tolerant infants showed a significant increase in fecal butyrate levels, and those taxa that were significantly enriched in these samples, Blautia and Roseburia, exhibited specific strain-level demarcations between tolerant and allergic infants. Our data suggest that EHCF+LGG promotes tolerance in infants with CMA, in part, by influencing the strain-level bacterial community structure of the infant gut.

Transplantation is the only cure for end-stage organ failure, but without immunosuppression, T cells rapidly reject allografts. While genetic disparities between donor and recipient are major determinants of the kinetics of transplant rejection, little is known about the contribution of environmental factors. Because colonized organs have worse transplant outcome than sterile organs, we tested the influence of host and donor microbiota on skin transplant rejection. Compared with untreated conventional mice, pretreatment of donors and recipients with broad-spectrum antibiotics (Abx) or use of germ-free (GF) donors and recipients resulted in prolonged survival of minor antigen-mismatched skin grafts. Increased graft survival correlated with reduced type I IFN signaling in antigen-presenting cells (APCs) and decreased priming of alloreactive T cells. Colonization of GF mice with fecal material from untreated conventional mice, but not from Abx-pretreated mice, enhanced the ability of APCs to prime alloreactive T cells and accelerated graft rejection, suggesting that alloimmunity is modulated by the composition of microbiota rather than the quantity of bacteria. Abx pretreatment of conventional mice also delayed rejection of major antigen-mismatched skin and MHC class II-mismatched cardiac allografts. This study demonstrates that Abx pretreatment prolongs graft survival, suggesting that targeting microbial constituents is a potential therapeutic strategy for enhancing graft acceptance.

SummaryBackground The role of the fungal microbiota in digestive diseases is poorly defined, but is becoming better understood due to advances in metagenomics. Aim To review the gastrointestinal fungal microbiota and its relationship with digestive diseases. Methods Search of the literature using PubMed and MEDLINE databases. Subject headings including ‘fungal-bacterial interactions’, ‘mycotoxins’, ‘immunity to fungi’, ‘fungal infection’, ‘fungal microbiota’, ‘mycobiome’ and ‘digestive diseases’ were used. Results The fungal microbiota is an integral part of the gastrointestinal microecosystem with up to 106 microorganisms per gram of faeces. Next-generation sequencing of the fungal 18S rRNA gene has allowed better characterisation of the gastrointestinal mycobiome. Numerous interactions between fungi and bacteria and the complex immune response to gastrointestinal commensal or pathogenic fungi all impact on the pathophysiology of inflammatory bowel disease and other gastrointestinal inflammatory entities such as peptic ulcers. Mycotoxins generated as fungal metabolites contribute to disturbances of gastrointestinal barrier and immune functions and are associated with chronic intestinal inflammatory conditions as well as hepatocellular and oesophagogastric cancer. Systemic and gastrointestinal disease can also lead to secondary fungal infections. Fungal genomic databases and methodologies need to be further developed and will allow a much better understanding of the diversity and function of the mycobiome in gastrointestinal inflammation, tumourigenesis, liver cirrhosis and transplantation, and its alteration as a consequence of antibiotic therapy and chemotherapy. Conclusions The fungal microbiota and its metabolites impact gastrointestinal function and contribute to the pathogenesis of digestive diseases. Further metagenomic analyses of the gastrointestinal mycobiome in health and disease is needed.

Currently considered incurable, multiple myeloma (MM) is characterized by proliferation of malignant plasma cells (PC) predominantly in the bone marrow, which overproduce monoclonal immunoglobulin proteins, and a perturbed tumor microenvironment, which promotes PC survival, inhibits osteoblast activity, increases osteoclast activity, and leads to hallmark osteolytic bone disease, all of which contribute to the clinical manifestations of the disease. Disrupting these sequelae by incorporation of proteasome inhibitors and immunomodulatory drugs into treatment regimens has improved overall survival of MM patients; however, the majority of patients will become refractory to the most effective currently available therapeutic options and relapse.1 It is hypothesized that a drug-resistant population of myeloma stem cells promotes disease relapse, but the characteristics and identity of this cell type(s) remain uncertain.2 Expression of maternal embryonic leucine zipper kinase (MELK) is increased in a number of cancers and is associated with poorer prognosis. MELK activity modulates many cellular and biological processes, including proliferation, apoptosis, hematopoiesis and oncogenesis, and is believed to have a critical role in cancer stem cell maintenance.3

The incidence of food allergy in developed countries is rising at a rate that cannot be attributed to genetic variation alone. In this review, we discuss the environmental factors that may contribute to the increasing prevalence of potentially fatal anaphylactic responses to food. Decreased exposure to enteric infections due to advances in vaccination and sanitation, along with the adoption of high-fat (Western) diets, antibiotic use, Cesarean birth, and formula feeding of infants, have all been implicated in altering the enteric microbiome away from its ancestral state. This collection of resident commensal microbes performs many important physiological functions and plays a central role in the development of the immune system. We hypothesize that alterations in the microbiome interfere with immune system maturation, resulting in impairment of IgA production, reduced abundance of regulatory T cells, and Th2-skewing of baseline immune responses which drive aberrant responses to innocuous (food) antigens.

Background & Aims The commensal microbiota is believed to have an important role in regulating immune responsiveness and preventing intestinal inflammation. Intestinal microbes produce signals that regulate inflammation via Toll-like receptor (TLR) signaling, but the mechanisms of this process are poorly understood. We investigated the role of the anti-inflammatory cytokine interleukin (IL)-10 in this signaling pathway using a mouse model of colitis. Methods Clinical, histopathologic, and functional parameters of intestinal inflammation were evaluated in TLR4 −/− , IL-10 −/− , and TLR4 −/− × IL-10 −/− mice that were free of specific pathogens and in TLR4 −/− × IL-10 −/− mice following eradication and reintroduction of Helicobacter hepaticus. Regulatory T-cell (Treg) function was evaluated by crossing each of the lines with transgenic mice that express green fluorescent protein under control of the endogenous regulatory elements of Foxp3 . Apoptotic cells in the colonic lamina propria were detected by a TUNEL assay. Results TLR4-mediated signals have 2 interrelated roles in promoting inflammation in TLR4 −/− × IL-10 −/− mice. In the absence of TLR4-mediated signals, secretion of proinflammatory and immunoregulatory cytokines is dysregulated. Tregs (Foxp3 + ) that secrete interferon-γ and IL-17 accumulate in the colonic lamina propria of TLR4 −/− × IL-10 −/− mice and do not prevent inflammation. Aberrant control of epithelial cell turnover results in the persistence of antigen-presenting cells that contain apoptotic epithelial fragments in the colonic lamina propria of Helicobacter- infected TLR4 −/− mice. Conclusions In mice that lack both IL-10- and TLR4-mediated signals, aberrant regulatory T-cell function and dysregulated control of epithelial homeostasis combine to exacerbate intestinal inflammation.